Showing posts with label development. Show all posts
Showing posts with label development. Show all posts

Thursday, 7 May 2015

Master Orchestrator of the Genome Is Discovered

Master Orchestrator of the Genome Is Discovered
Thursday, 07 May 2015

One of developmental biology’s most perplexing questions concerns what signals transform masses of undifferentiated cells into tremendously complex organisms, a process called ontogeny.

UB research suggests a new paradigm, visualized
in this diagram, for developmental global genome
programming by the nuclear FGFR1 protein.
New research by University at Buffalo scientists, published last week in PLOS ONE, provides evidence that it all begins with a single “master” growth factor receptor that regulates the entire genome.

“The finding provides a new level of understanding of the fundamental aspects of how organisms develop,” says senior author Michal K. Stachowiak, PhD, professor in the Department of Pathology and Anatomical Sciences in the UB School of Medicine and Biomedical Sciences and senior author. He also directs the Stem Cell Engraftment and In Vivo Analysis Facility and the Stem Cell Culture and Training Facility at the Western New York Stem Cell Culture and Analysis Center at UB.

“Our research shows how a single growth factor receptor protein moves directly to the nucleus in order to program the entire genome,” he said.

Michal Stachowiak, PhD, Professor, Department
of Pathology and Anatomical Sciences. 
The research challenges a long-held supposition in biology that specific types of growth factors only functioned at a cell’s surface. For two decades, Stachowiak’s team has been intrigued by the possibility that growth factors function from within the nucleus, a point, he says, this current paper finally proves.

A more advanced understanding of how organisms form, based on this work, has the potential to significantly enhance the understanding and treatment of cancers, which result from uncontrolled development as well as congenital diseases, the researchers say. The new research also will contribute to the understanding of how stem cells work.

This work was conducted on mouse embryonic stem cells, not human cells.

Organizing ‘this cacophony of genes’
“We’ve known that the human body has almost 30,000 genes that must be controlled by thousands of transcription factors that bind to those genes,” Stachowiak said, “yet we didn’t understand how the activities of genes were coordinated so that they properly develop into an organism.”

“Now we think we have discovered what may be the most important player, which organizes this cacophony of genes into a symphony of biological development with logical pathways and circuits,” he said.

At the centre of the discovery is a single protein called nuclear Fibroblast Growth Factor Receptor 1 (nFGFR1).

“FGFR1 occupies a position at the top of the gene hierarchy that directs the development of multicellular animals,” said Stachowiak.

The FGFR1 gene is known to govern gastrulation, occurring in early development, where the three-layered embryonic structure forms. It also plays a major role in the development of the central and peripheral nervous systems and the development of the body’s major systems, including muscles and bones.

To study how nuclear FGFR1 worked, the UB team used genome-wide sequencing of mouse embryonic stem cells programmed to develop cells of the nervous system, with additional experiments in which nuclear FGFR1 was either introduced or blocked. The researchers found that the protein was responsible, either alone or with so-called partner nuclear receptors, for ensuring that embryonic stem cells develop into differentiated cells. By targeting thousands of genes, it controls the development of the major points of growth in the body (known as axes) as well as neuronal and muscle development.

The research shows that nuclear FGFR1 binds to promoters of genes that encode transcription factors, the proteins that control which genes are turned on or off in the genome.

“We found that this protein works as a kind of ‘orchestration factor,’ preferably targeting certain gene promoters and enhancers. The idea that a single protein could bind thousands of genes and then organize them into a hierarchy, that was unknown,” Stachowiak said.

“Nobody predicted it.”

Sequencing advances
The discovery that a single protein can exert such a global genomic function stems from recent advances in DNA sequencing technologies, which allow for the sequencing of a complex genome in just hours.

“NextGen DNA sequencing allows us to analyse millions of DNA sequences selected by the interacting protein,” Stachowiak said.

In the UB research, the DNA sequencing data were processed by the supercomputer at the university’s Center for Computational Research (CCR). Stachowiak and his colleagues then spent weeks aligning these data to the genome and conducting further analyses.

“We imposed nuclear FGFR1 on every little corner of genome,” he said.

“The computer spit out which genes are affected by nuclear FGFR1: it was an enormously complex network of genome activity.”

They found that the protein binds to genes that make neurons and muscles as well as to an important oncogene, TP63, which is involved in a number of common cancers.

Other studies in Stachowiak’s laboratory demonstrate that these interactions also take place in the human genome, controlling function and possibly underlying diseases like schizophrenia. Targeting of the nuclear FGFR1 allows for the reactivation of neural development in the adult brain in preclinical studies and thus, Stachowiak says, may offer unprecedented opportunity for regenerative medicine. Nuclear accumulation of nuclear FGFR1 may be altered in some cancer cells, and thus could become a focus in cancer therapy, he added.  

“This seminal discovery lends new perspectives to the origin, nature and treatment of a variety of human disease,” Stachowiak concluded.

Source: University at Buffalo, New York
Contact: Ellen Goldbaum

Reference:
Global Developmental Gene Programing Involves a Nuclear Form of Fibroblast Growth Factor Receptor-1 (FGFR1)
Christopher Terranova,Sridhar T. Narla, Yu-Wei Lee, Jonathan Bard, Abhirath Parikh, Ewa K. Stachowiak, Emmanuel S. Tzanakakis, Michael J. Buck, Barbara Birkaya, Michal K. Stachowiak
PLoS ONE 2015 10(4):e0123380, doi:10.1371/journal.pone.0123380
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For more on stem cells and cloning, go to CellNEWS at

New Stem Cell May Overcome Hurdles for Regenerative Medicine

Salk Institute scientists discover new type of stem cell that could potentially generate mature, functional tissues
Thursday, 07 May 2015

In this image, a novel type of human stem cell is
shown in green integrating and developing into
the surrounding cells of a nonviable mouse
embryo. Red indicates cells of endoderm lineage.
Endoderm cells can give rise to tissue that covers
organs from the digestive and respiratory
systems. The new stem cell, developed at the
Salk Institute, holds promise for one day growing
replacement functional cells and tissues. Credit:
Courtesy of the Salk Institute for Biological

Studies.
Scientists at the Salk Institute have discovered a novel type of pluripotent stem cell – cells capable of developing into any type of tissue – whose identity is tied to their location in a developing embryo. This contrasts with stem cells traditionally used in scientific study, which are characterized by their time-related stage of development.

In the paper, published May 6, 2015 in Nature, the scientists report using these new stem cells to develop the first reliable method for integrating human stem cells into nonviable mouse embryos in a laboratory dish in such a way that the human cells began to differentiate into early-stage tissues.

“The region-specific cells we found could provide tremendous advantages in the laboratory to study development, evolution and disease, and may offer avenues for generating novel therapies,” says Salk Professor Juan Carlos Izpisua Belmonte, senior author of the paper and holder of Salk’s Roger Guillemin Chair.

The researchers dubbed this new class of cells “region-selective pluripotent stem cells,” or rsPSCs for short. The rsPSCs were easier to grow in the laboratory than conventional human pluripotent stem cells and offered advantages for large-scale production and gene editing (altering a cell’s DNA), both desirable features for cell replacement therapies.

Juan Carlos Izpisua Belmonte and Jun Wu
Credit: Courtesy of the Salk Institute for
Biological Studies.
To produce the cells, the Salk scientists developed a combination of chemical signals that directed human stem cells in a laboratory dish to become spatially oriented.

They then inserted the spatially oriented human stem cells (human rsPSCs) into specific regions of partially dissected mouse embryos and cultured them in a dish for 36 hours. Separately, they also inserted human stem cells cultured using conventional methods, so that they could compare existing techniques to their new technique.

While the human stem cells derived through conventional methods failed to integrate into the modified embryos, the human rsPSCs began to develop into early stage tissues. The cells in this region of an early embryo undergo dynamic changes to give rise to all cells, tissues and organs of the body. Indeed the human rsPSCs began the process of differentiating into the three major cell layers in early development, known as ectoderm, mesoderm and endoderm. The Salk researchers stopped the cells from differentiating further, but each germ layer was theoretically capable of giving rise to specific tissues and organs.

The new stem cell (green), developed at the Salk
Institute, holds promise for one day growing
replacement functional cells and tissues. Credit:
Courtesy of the Salk Institute for Biological

Studies.
Collaborating with the labs of Salk Professors Joseph Ecker and Alan Saghatelian, the Izpisua Belmonte team performed extensive characterization of the new cells and found rsPSCs showed distinct molecular and metabolic characteristics as well as novel epigenetic signatures – that is, patterns of chemical modifications to DNA that control which genes are turned on or off without changing the DNA sequence.

“The region selective-state of these stem cells is entirely novel for laboratory-cultured stem cells and offers important insight into how human stem cells might be differentiated into derivatives that give rise to a wide range of tissues and organs,” says Jun Wu, a postdoctoral researcher in Izpisua Belmonte’s lab and first author of the new paper.

“Not only do we need to consider the timing, but also the spatial characteristics of the stem cells. Understanding both aspects of a stem cell’s identity could be crucial to generate functional and mature cell types for regenerative medicine.”

Contact: Salk Communications

Reference:
An alternative pluripotent state confers interspecies chimaeric competency
Authors: Jun Wu, Daiji Okamura, Mo Li, Keiichiro Suzuki, Chongyuan Luo,Li Ma, Yupeng He, Zhongwei Li, Chris Benner, Isao Tamura, Marie N. Krause, Joseph R. Nery, Tingting Du, Zhuzhu Zhang, Tomoaki Hishida, Yuta Takahashi, Emi Aizawa, Na Young Kim, Jeronimo Lajara, Pedro Guillen, Josep M. Campistol, Concepcion Rodriguez Esteban, Pablo J. Ross, Alan Saghatelian, Bing Ren, Joseph R. Ecker and Juan Carlos Izpisua Belmonte
Nature, 06 May 2015, doi:10.1038/nature14413
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For more on stem cells and cloning, go to CellNEWS at
http://cellnews-blog.blogspot.com/

Friday, 30 January 2015

Why is a Dolphin Not a Cat?

How repurposing non-coding elements in the genome gave rise to the great 'mammalian radiation'
Friday, 30 January 2015

Exploring gene regulation in 20 mammals
provides insights into the mammalian radiation
that occurred over 100 million years ago. Credit:
Spencer Phillips, EMBL-EBI.
New research shows how evolution has given rise to a rich diversity of species by repurposing functional elements shared by all mammals. Published in Cell by scientists at the European Bioinformatics Institute (EMBL-EBI) and the University of Cambridge Cancer Research UK-Cambridge Institute (CRUK CI), the study demonstrates how methods for understanding human biology can be used to understand a broad range of species.

Mammals all share a common ancestor, and they share a lot of the same genes. So what exactly makes a dolphin not a cat, and how did we all start to diverge from one another millions of years ago? Part of the answer lies in how - and when - genes are regulated. This latest research explores the evolution of gene regulation in 20 mammalian species, and provides deep insights into the 'mammalian radiation', a time of rapid morphological evolution that occurred shortly after the asteroid impact that caused the extinction of the dinosaurs.

Leveraging findings from a study comparing the genome sequences of 29 mammals, and with the help of conservation organisations such as the UK Cetacean Strandings Investigation Programme and the Copenhagen Zoo, the team were able to study and compare gene regulation in liver cells from 20 key species including the naked mole rat, human, Tasmanian devil, dolphin and Sei whale.

"What we've shown is that evolution repurposes things that exist in all species, to make each species unique," explains Paul Flicek, head of Vertebrate Genomics at EMBL-EBI.

"By looking at gene promoters and enhancers in many different mammals, we demonstrated that species-specific enhancers come from ancient DNA - that evolution captures DNA that's been around for a long time, and uses it for gene regulation in specific tissues."

Evolution has two ways to turn changes in the genome into differences between species: it can change a protein sequence, or it can change the way promoters or enhancers control that protein's expression. Today's study also shows that in some cases evolution uses both strategies at once. When amino acid sequences evolve very quickly, important regulation changes occur at the same time: the protein-coding sequence and the corresponding regulatory sequence change synergistically.

Gathering the samples - the experimental efforts were led by Diego Villar of CRUK CI - took well over two years, and the experiments themselves produced a staggering volume of data. Analysing the results brought the team to a new frontier in bioinformatics.

"People spend a lot of time and money trying to understand human biology, so most of the tools we have are designed to study human genomes," explains Camille Berthelot of EMBL-EBI, who led the computational work.

"The reference data we have for the less studied species, like the Sei whale or Tasmanian devil, are nothing like the pored-over datasets we have for the human genome. A lot of what we did involved benchmarking, and making sure the methods and algorithms were fit for this kind of comparison."

"What inspired this work was a desire to get on top of the mountain, look out and see what is going on in the landscape of molecular evolution across the breadth of mammalian space," says Duncan Odom of CRUK CI and Wellcome Trust Sanger Institute.

"What's exciting about this study is that we now know we can start to answer questions about the functional genetics of many under-explored species - questions we usually can ask only of humans and mice. We can use tools developed to study humans to understand the biology of all kinds of animals, whether they're blackbirds or elephants, and explore their relationship with one another. This research has given us new insights into mammalian evolution, and proven how powerful these methods can be."

Source: EMBL
Contact: Mary Todd Bergman

Reference:
Enhancer Evolution across 20 Mammalian Species
Diego Villar, Camille Berthelot, Sarah Aldridge, Tim F. Rayner, Margus Lukk, Miguel Pignatelli, Thomas J. Park, Robert Deaville, Jonathan T. Erichsen, Anna J. Jasinska, James M.A. Turner, Mads F. Bertelsen, Elizabeth P. Murchison, Paul Flicek, Duncan T. Odom
Cell, Volume 160, Issue 3, p554–566, 29 January 2015, DOI: http://dx.doi.org/10.1016/j.cell.2015.01.006
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For more on stem cells and cloning, go to CellNEWS at

Tuesday, 4 November 2014

Researchers Reconstruct Early Stages of Embryo Development

Researchers Reconstruct Early Stages of Embryo Development
Tuesday, 04 November 2014

Researchers at the University of Cambridge have managed to reconstruct the early stage of mammalian development using embryonic stem cells, showing that a critical mass of cells – not too few, but not too many – is needed for the cells to being self-organising into the correct structure for an embryo to form.


All organisms develop from embryos: a cell divides generating many cells. In the early stages of this process, all cells look alike and tend to aggregate into a featureless structure, more often than not a ball. Then, the cells begin to 'specialise' into different types of cell and space out asymmetrically, forming an axis which begins to provide a structure for the embryo to develop along.

In animal embryos this stage is followed by a process known as gastrulation: a choreographed movement of the cells that, using the initial axis as a reference, positions the head and the tail, the front and the back. During the process, the cells begin to form three distinct layers: the endoderm, mesoderm and ectoderm, determining which tissues or organs the cells will then develop into.

Professor Alfonso Martinez-Arias from the Department of Genetics at the University of Cambridge, who led the research, says:

"Gastrulation was described by biologist Professor Lewis Wolpert as being 'truly the most important event in your life' because it creates the blueprint of an organism. Axis formation and gastrulation are the two central processes that initiate the development of an organism and are inextricably associated with the embryo. We have managed to recreate this for the first time in the lab."

Professor Martinez-Arias and colleagues, supported by the European Research Council and the Wellcome Trust, have reconstructed these early stages of development using mouse embryonic stem cells. Embryonic stem cells, discovered in the Department of Genetics in the 1980s (for which Sir Martin Evans was awarded the Nobel Prize in Physiology or Medicine 2007), have become an important tool for developmental biology, understanding disease, and in regenerative medicine due to the ability to give rise to all cell types in culture. Over the last few years, they have been used to 'grow' organs including the eye and the cerebral cortex; surprisingly, these structures develop without an axis.

In research published today in the journal Development, the researchers report a way to coax cells to reorganize in the manner that they do in an embryo, creating an axis and undergoing movements and organisations that mimic the process of gastrulation. Over the years researchers have been making aggregates of embryonic stem cells to obtain certain cell types, for example red blood cells. However, these aggregates lack structure and the different cell types emerge in a disorganised fashion. This is the first time that researchers have been able to elicit axis formation, spatial organisation and gastrulation-like movements from aggregates of embryonic stem cells.

The researchers show that if the number of cells aggregated initially is similar to that of a mouse embryo, the cells generate a single axis and this serves as a template for a sequence of events that mimics those of the early embryo. By manipulating the signals that the cells see at a particular time, the researchers were able to influence what type of cell they become and how they are organised. In one of the experiments, for example, activation of a particular signal at the correct time elicits the appearance of the mesoderm, endoderm and ectoderm – the precursors of all cell types – with a spatial organization similar to that of an embryo.

Using this experimental system, the researchers were able to generate the early stages of a spinal cord, which they showed forms as part of the process of gastrulation. This finding complements previous research from the University of Edinburgh and the National Institute for Medical Research which showed that embryonic stem cells can be coaxed into this spinal cord cells; however, the Cambridge researchers showed that in the embryo-like aggregates, the structural organization is more robust and allows for the polarised growth of the tissue.

Professor Martinez-Arias adds:

"It is early days but this system promises insights into the early stages of development and what determines the specification of the different cell types. This will allow more robust protocols for differentiation with cues that mimic those that the cells are subject to in embryos.”

"Most significantly, the system will provide a means to test, experimentally, how a homogeneous group of cells organizes itself in space, a central process in the development of any organism, and the ability to recreate in culture the niches that adult stem cells create during embryogenesis and which have remained elusive experimentally."

Contact: Craig Brierley

References:
Symmetry breaking, germ layer specification and axial organisation in aggregates of mouse ES cells
Susanne C. van den Brink, Peter Baillie-Johnson, Tina Balayo, Anna-Katerina Hadjantonakis, Sonja Nowotschin, David A. Turner, and Alfonso Martinez Arias
Development 2014 141:4231-4242; doi:10.1242/dev.113001

Wnt/β-catenin and FGF signalling direct the specification and maintenance of a neuromesodermal axial progenitor in ensembles of mouse ES cells
David A. Turner, Penelope C. Hayward, Peter Baillie-Johnson, Pau Rué, Rebecca Broome, Fernando Faunes, and Alfonso Martinez Arias
Development; 2014 141:4243-4253; doi:10.1242/dev.112979
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For more on stem cells and cloning, go to CellNEWS at

Tuesday, 1 July 2014

Researchers Coax Human Embryonic Stem Cells to Organize Themselves Using Geometry

Using Geometry, Researchers Coax Human Embryonic Stem Cells to Organize Themselves
Tuesday, 01 July 2014

About seven days after conception, something remarkable occurs in the clump of cells that will eventually become a new human being. They start to specialize. They take on characteristics that begin to hint at their ultimate fate as part of the skin, brain, muscle or any of the roughly 200 cell types that exist in people, and they start to form distinct layers.

Forty-two hours after they began to differentiate,
embryonic cells are clearly segregating into
endoderm (red), mesoderm (blue) and ectoderm
(black). Researchers say the key to achieving this
patterning in culture is confining the colonies
geometrically. Credit: Brivanlou Lab, the
Rockefeller University. 
Although scientists have studied this process in animals, and have tried to coax human embryonic stem cells into taking shape by flooding them with chemical signals, until now the process has not been successfully replicated in the lab. But researchers led by Ali Brivanlou, Robert and Harriet Heilbrunn Professor and head of the Laboratory of Stem Cell Biology and Molecular Embryology at The Rockefeller University, have done it, and it turns out that the missing ingredient is geometrical, not chemical.

"Understanding what happens in this moment, when individual members of this mass of embryonic stem cells begin to specialize for the very first time and organize themselves into layers, will be a key to harnessing the promise of regenerative medicine," Brivanlou says.

"It brings us closer to the possibility of replacement organs grown in petri dishes and wounds that can be swiftly healed."

In the uterus, human embryonic stem cells receive chemical cues from the surrounding tissue that signal them to begin forming layers — a process called gastrulation. Cells in the centre begin to form ectoderm, the brain and skin of the embryo, while those migrating to the outside become mesoderm and endoderm, destined to become muscle and blood and many of the major organs, respectively.

Brivanlou and his colleagues, including postdocs Aryeh Warmflash and Benoit Sorre as well as Eric Siggia, Viola Ward Brinning and Elbert Calhoun Brinning Professor and head of the Laboratory of Theoretical Condensed Matter Physics, confined human embryonic stem cells originally derived at Rockefeller to tiny circular patterns on glass plates that had been chemically treated to form "micropatterns" that prevent the colonies from expanding outside a specific radius. When the researchers introduced chemical signals spurring the cells to begin gastrulation, they found the colonies that were geometrically confined in this way proceeded to form endoderm, mesoderm and ectoderm and began to organize themselves just as they would have under natural conditions. Cells that were not confined did not.

By monitoring specific molecular pathways the human cells use to communicate with one another to form patterns during gastrulation — something that was not previously possible because of the lack of a suitable laboratory model — the researchers also learned how specific inhibitory signals generated in response to the initial chemical cues function to prevent the cells within a colony from all following the same developmental path.

"At the fundamental level, what we have developed is a new model to explore how human embryonic stem cells first differentiate into separate populations with a very reproducible spatial order just as in an embryo," says Warmflash.

"We can now follow individual cells in real time in order to find out what makes them specialize, and we can begin to ask questions about the underlying genetics of this process."

The research also has direct implications for biologists working to create "pure" populations of specific cells, or engineered tissues consisting of multiple cell types, for use in medical treatments.

"These cells have a powerful intrinsic tendency to form patterns as they develop," Warmflash says.

"Varying the geometry of the colonies may turn out to be an important tool that can be used to guide stem cells to form specific cell types or tissues."

Contact: Zach Veilleux

Reference:
A method to recapitulate early embryonic spatial patterning in human embryonic stem cells
Aryeh Warmflash, Benoit Sorre, Fred Etoc, Eric D. Siggia and Ali H. Brivanlou
Nature Methods online: June 29, 2014, doi:10.1038/nmeth.3016
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